1887

Abstract

Summary: The activity of the proline catabolic enzyme pyrroline-5-carboxylate dehydrogenase (EC 1.5.1.12) was induced up to threehundredfold by the addition of three hundred proline to the growth medium of the Gram-positive bacterium Streptomyces coelicolor A3(2). Rifampicin, an inhibitor of RNA polymerase activity, abolished induction, implying that regulation was at the level of activation of gene transcription. The enzyme was purified and SDS-PAGE of the highly purified enzyme preparation revealed a single subunit with 68000. A single band of protein, which also stained for enzyme activity, was observed after native gel electrophoresis. The of the enzyme was estimated to be approximately 265000 by native gel electrophoresis and approximately 305000 by gel filtration, which indicated that the enzyme had a tetrameric quaternary structure. The apparent for pyrroline-5-carboxylate was 109 pm7-3 M, whilst that for NAD was 43-3+2-5 M. Product inhibition by NADH (apparent 0-6 mM) was observed. The observed was 22-0 pm 1 mol min (mg protein). Neither 1 nor 5 mM proline had any effect on enzyme activity, whilst glutamate was a very weak inhibitor.

Loading

Article metrics loading...

/content/journal/micro/10.1099/13500872-141-7-1739
1995-07-01
2024-04-20
Loading full text...

Full text loading...

/deliver/fulltext/micro/141/7/mic-141-7-1739.html?itemId=/content/journal/micro/10.1099/13500872-141-7-1739&mimeType=html&fmt=ahah

References

  1. Farres J., Julia P., Pares X. 1988; Aldehyde oxidation in human placenta: purification and properties of l-pyrroline-5- carboxylate dehydrogenase.. Biochem J 256:461–467
    [Google Scholar]
  2. Hodgson D.A. 1982; Glucose repression of carbon source uptake and metabolism in Streptomyces coelicolor A3(2) and its perturbation in mutants resistant to 2-deoxyglucose.. J Gen Microbiol 128:2417–2430
    [Google Scholar]
  3. Hood D.W., Heidstra R., Swoboda U.K., Hodgson D.A. 1992; Molecular genetic analysis of proline and tryptophan biosynthesis in Streptomyces coelicolor A3(2): interaction between primary and secondary metabolism - a review.. Gene 115:5–12
    [Google Scholar]
  4. Hopwood D.A., Bibb M.J., Chater K.F., Kieser T., Bruton C.P., Kieser H.M., Lydiate D.J., Smith C.P., Ward J.M., Schrempf H. 1985 Genetic Manipulation of Streptomyces-. a Laboratory Manual. Norwich: The John Innes Foundation;
    [Google Scholar]
  5. Isobe K., Matsuzawa T., Soda K. 1987; Crystallization and characterization of the l-pyrroline-5-carboxylate dehydrogenase from Bacillus sphaericus.. Agric Biol Chem 51:1947–1953
    [Google Scholar]
  6. Kieser H.M., Kieser T., Hopwood D.A. 1992; A combined genetic and physical map of the Streptomyces coelicolor A3(2) chromosome.. J Bacteriol 174:5496–5507
    [Google Scholar]
  7. Meile L., Leisinger T. 1982; Purification and properties of the bifunctional proline dehydrogenase l-pyrroline-5-carboxylate dehydrogenase from Pseudomonas aeruginosa.. Eur J Biochem 129:67–75
    [Google Scholar]
  8. Menzel R., Roth J. 1981; Purification of the put A gene product: a bifunctional, membrane bound protein from Salmonella typhimurium responsible for the 2-step oxidation of proline to glutamate.. J Biol Chem 256:9755–9761
    [Google Scholar]
  9. Small W.C., Jones M.E. 1990; Pyrroline-5 -carboxylate dehydrogenase of the mitochondrial matrix of rat liver: purification, physical and kinetic characteristics.. J Biol Chem 265:18668–18672
    [Google Scholar]
  10. Strauch E., Takano E., BayIis H.A., Bibb M.J. 1991; Thestringent response in Streptomyces coelicolor A3(2).. Mol Microbiol 5:289–298
    [Google Scholar]
  11. Tsuji R.F., Magae J., Yamashita M., Nagai K., Yamasaki M. 1992; Selective immunosuppression of prodigiosin 25C and FK506 in the murine immune system.. J Antibiot 45:1295–1302
    [Google Scholar]
  12. Wasserman H.H., Shaw C.K., Sykes R.J., Cushley R.J. 1974; The biosynthesis of metacycloprodigiosin and undecyl- prodigiosin.. Tetrahedron Lett 33:2787–2790
    [Google Scholar]
  13. Williams I., Frank L. 1975; Improved chemical synthesis and enzymatic assay of A-l-pyrroline-5-carboxylic acid.. Anal Biochem 64:85–97
    [Google Scholar]
http://instance.metastore.ingenta.com/content/journal/micro/10.1099/13500872-141-7-1739
Loading
/content/journal/micro/10.1099/13500872-141-7-1739
Loading

Data & Media loading...

This is a required field
Please enter a valid email address
Approval was a Success
Invalid data
An Error Occurred
Approval was partially successful, following selected items could not be processed due to error